This invention relates generally to a device, and process, for continuously separating solutes and suspended particles from a fluid phase in a magnetically stabilized fluidized bed (MSFB), and more particularly, to a device and process wherein multiple solutes in a fluidizing medium can be retained and concentrated against the action of diffusion and dispersion (i.e., "focused") at specific respective locations within an MSFB by adjusting the binding equilibrium throughout the bed by imposing a stationary property gradient, such as a pH gradient, on the moving bed.
There is a need, particularly in the biotechnology field, for improved product recovery techniques. The separation and purification of product from a typical biochemical reactor can require extensive capital, space, and operation coats. Indeed, separation processes frequently comprise a substantial proportion of the product's cost. Ideally, separation processes should be inexpensive, capable of handling large volumes of liquids and cells or cell debris, and efficient. Conventional techniques for biochemical product recovery typically include filtration or centrifugation to remove any suspended solids followed by a batch separation operation, and, in some cases, an additional purification operation(s). Conventional separation and purification operations are typically performed in chromatography and adsorption (e.g., ion-exchange or affinity) packed columns.
Isoelectric focusing is a known technique for continuously separating solutes from an aqueous solution based on the isoelectric point of the components. In an exemplary process, a mixture of protein molecules is resolved into its components by subjecting the mixture to an electric field in a supporting gel having a previously established pH gradient. The isoelectric point, pI, is the pH value at which the net charge on a molecule in solution is zero. At pI, the molecule will not move in an electric field. The isoelectric focusing technique can be used to focus proteins and amino acids whose pI points differ by less than 0.02 pH units.
The isoelectric focusing technique, however, requires special chemicals, ampholytes, to create the pH gradient and the application of a high-power electric field. The ampholytes must be specially synthesized thereby adding expense to the system. Disadvantageously, the separated solutes may contain contaminants from the ampholytes and other solutes within the aqueous solution. This necessitates at least one additional step to separate the solute from the ampholyte.
An isoelectric focusing device has been devised to obviate the need for ampholytes by using a gel pore size gradient to effect separation. This device also requires an electric field for partial or total solute mobility.
The current used to generate the field in isoelectric focusing systems can create convective mixing. Further, the electric field generates he, at which can adversely affect heat-sensitive solutes. In many cases, a cooling system is required which further increases the complexity and cost of the isoelectric focusing system.
Thus, while isoelectric focusing may produce concentrated solute samples, the high power requirements and heat generation associated with the use of electric fields are major drawbacks. A second technique, chromatofocusing, eliminates the need for the electric field, but suffers from the inability to run continuously or at steady-state. Chromatofocusing systems, however, use simple ion-exchange resins and buffers thereby obviating the need for ampholytes and the complications attendant thereto.
The chromatofocusing technique also separates solutes on the basis of pI. An aqueous solution, buffered to a predetermined pH is introduced into a chromatography column containing an ion-exchange resin at a different pH. As the solution flows through the column, a continually changing pH gradient is formed throughout the column. The pH gradient changes from a value near the exchange resin pit to a near linear gradient between the solution buffer and the exchange resin pH to a final pH equal to the solution buffer. Since both the solution and the exchange resin have some buffering capacity, the movement of any particular pH value through the column is much slower than the movement of the liquid solution phase. This differential motion can be used to focus solutes in the column.
Illustratively, the liquid buffer solution has a very high pH, the column is initially at a very low pH, and the solid ion exchange resin is negatively charged. Solutes introduced with the liquid buffer solution would be negatively charged, and therefore, would be repelled by the ion exchange resin. As the solutes travel down the column, the pH values become lower. Eventually, each solute will reach a region where the pH equals its pI. Beyond this point, the solute will change charge and adsorb on the resin. In this manner, each solute will focus at its particular pI value, slowly move down the column at the velocity of the pH gradient, and elute from the column in a focused pulse at its pI.
For focusing particulates, such as cells, packed columns tend to clog at high cell concentrations. Isoelectric focusing can separate cells, but has the drawbacks of ohmeric heating and high power consumption. A further disadvantage of chromatofocusing is that it is inherently a batch-wise operation. Typically, the efficiency of batch operations is limited by column height and by the necessity of regenerating the adsorbent for use in subsequent operations. All of the foregoing disadvantages are serious drawbacks for large scale commercial operations.
Continuous fixed bed processes have been proposed to compensate for the disadvantages of a stationary column, batch process. However, continuous fixed beds are mechanically complicated and the efficiency is limited by particle size of the adsorbent. Smaller adsorbent particles give better resolution, but result in a large pressure drop across the bed. The pressure drop can be reduced by fluidizing the bed. However, fluidized beds have the disadvantages of backmixing and channel formation.
Magnetically stabilized fluidized beds have been developed to compensate for the aforementioned disadvantages of a fluidized bed. A magnetically stabilized fluidized bed consists of a column or bed packed with solid support particles which are magnetically susceptible. A fluidizing medium, such as a liquid or gas, is introduced into the bed at a velocity sufficient to fluidize the solid particles, i.e., to cause the particles to rapidly mix and act as a fluid phase. The fluid properties of the solids permit them to flow out of the column for regeneration or replacement. Stabilization of the bed with a uniform low-power magnetic field (&lt;10 watts/liter) eliminates the mixing and turbulence effects otherwise observed in a fluidized bed. As soon as the magnetic field is applied, the solid particles stop mixing and remain motionless. The solid particles travel through the bed in smooth plug flow movement. The flow of fluid through a magnetically stabilized fluidized bed is similar to a conventional packed bed.
One drawback of magnetically stabilized fluidized beds is that the magnetically susceptible resins have to be specially synthesized or prepared. It has been discovered that many nonmagnetic resins, such as the ion-exchange resins sold under the trademarks Amberlite.RTM. (Rohm and Haas, Philadelphia, Pa.) or Dowex.RTM. (The Dow Chemical Company, Midland, Mich.) can be combined with magnetically susceptible solids such as nickel or stainless steel spheres to form a "mixed bed." Stable mixed beds have been formed containing up to about 80% nonmagnetic resin by volume.
Typically, magnetically stabilized fluidized bed processes are limited by the column height, which limits the amount of adsorbent available for separations, and the requirement of a specific adsorbent-desorbent system.
One suggested solution, termed "isomobility focusing," is described in U.S. Pat. No. 4,780,113 to Koslow. Isomobility focusing is defined as the process of selectively concentrating and separating a component of a mixture in a controllably transported MSFB as a result of controlling conditions of the bed such that the movement of a given component of the feedstream in the fluid phase is equal to the movement of that component on the solid phase. If the two phases are moving in opposite directions, the component will have no net velocity, its movement in one phase being equal to its movement in the other phase. Various methods for controlling the process conditions, such as adjusting the chemical or physical properties of the fluid phase, such as pH, temperature, pressure, or salt concentration are suggested. The control of process conditions is uniform over the entire isomobility focusing zone to adjust the affinity of the chemical specie being separated. Thus, isomobility focusing can not concentrate and/or separate multiple components on the same bed.
Further, although isomobility focusing is described using the term "focusing," the product is not concentrated at a given point along the column length, but rather is diluted within the column as the product bandwidth increases.
It is, therefore, an object of this invention to provide a device and process for continuously separating dissolved solutes and suspended particles from a multicomponent fluid phase in a magnetically stabilized fluidized bed.
It is another object of this invention to provide a device and process for continuously separating dissolved solutes and suspended particles in a magnetically stabilized fluidized bed with no cross-contamination of products and/or compounds used in the processing.
It is also an object of this invention to provide a device and process for continuously separating dissolved solutes and suspended particles in a magnetically stabilized fluidized bed using standard adsorption resins and buffers.
It is a further object of this invention to provide a device and process for continuously separating dissolved solutes and suspended particles in a magnetically stabilized fluidized bed without the need for specially prepared adsorbents or amphoteric compounds.
It is additionally an object of this invention to provide a device and process for continuously separating dissolved solutes and suspended particles in a magnetically stabilized fluidized bed without the use of a heat generating electric field.
It is yet a further object of this invention to provide a device and process for continuously separating dissolved solutes and suspended particles in a magnetically stabilized fluidized bed so that both solutes and suspended particles can be focused, i.e., increased to many times their original concentration.
It is also another object of this invention to provide a device and process for continuously separating dissolved solutes and suspended particles in a magnetically stabilized fluidized bed wherein separation time is fast (preferably on the order of minutes).
It is yet an additional object of this invention to provide a device and process for continuously separating dissolved solutes and suspended particles in a magnetically stabilized fluidized bed wherein operation, control and scale-up are simple and relatively inexpensive.